Estimation of interaction between oriented immobilized green fluorescent protein and its antibody by high performance affinity chromatography and molecular docking
Estimation of interaction between oriented immobilized green fluorescent protein and its antibody by high performance affinity chromatography and molecular docking
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通过高效亲和层析和分子对接评估定向固定化绿色荧光蛋白与其抗体之间的相互作用
DOI：
10.1002/jmr.2460
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发表时间：
2015-07-01
影响因子：
2.7
通讯作者：
Zheng, Xiaohui
中科院分区：
文献类型：
--
作者：
Li, Qian;Wang, Jing;Zheng, Xiaohui
尽管绿色荧光蛋白（GFP）及其抗体在生命科学中被广泛用于追踪蛋白质或细胞，但它们之间的结合行为仍不清楚。在这项工作中，通过重氮偶联法合成了一种新的固定化GFP，并将其定向固定在大孔硅胶表面。利用该固定相，在以GFP抗体为探针探索蛋白质 - 蛋白质相互作用时，证实了进样量依赖分析的有效性。结果证明GFP抗体在固定化的GFP上具有一种结合位点。结合位点的数量和结合常数经计算分别为(6.41 ± 0.76)×10⁻¹⁰ M和(1.39 ± 0.12)×10⁹ M⁻¹。通过分子对接进一步分析表明，GFP与其抗体的结合主要由氢键和盐桥驱动。这些结果表明，进样量依赖分析能够探索蛋白质 - 蛋白质相互作用，具有节省配体和时间的优势。对于昂贵或难以获得的配体来说，这是一种有价值的方法。版权所有（c）2015约翰威立父子有限公司
Although green fluorescence protein (GFP) and its antibody are widely used to track a protein or a cell in life sciences, the binding behavior between them remains unclear. In this work, diazo coupling method that synthesized a new stationary GFP was oriented immobilized on the surface of macro‐porous silica gel by a phase. The stationary phase was utilized to confirm the validation of injection amount‐dependent analysis in exploring protein–protein interaction that use GFP antibody as a probe. GFP antibody was proved to have one type of binding site on immobilized GFP. The number of binding site and association constant were calculated to be (6.41 ± 0.76) × 10‐10 M and (1.39 ± 0.12) × 109 M‐1. Further analysis by molecular docking showed that the binding of GFP to its antibody is mainly driven by hydrogen bonds and salt bridges. These results indicated that injection amount‐dependent analysis is capable of exploring the protein–protein interactions with the advantages of ligand and time saving. It is a valuable methodology for the ligands, which are expensive or difficult to obtain. Copyright © 2015 John Wiley & Sons, Ltd.